Combined effects of Allium sativum and Cuminum cyminum essential oils on planktonic and biofilm forms of Salmonella typhimurium isolates

Sa lmonella typhimurium (S. typhimurium) represents an important global public health problem and has the ability to survive under desiccation conditions in foods and food processing facilities for years. The aim of this study was to investigate the effects of Allium sativum (A. sativum) and Cuminum cyminum (C. cyminum) essential oils (EOs) against planktonic growth, biofilm formation and quorum sensing (QS) of S. Typhimurium isolates, the strong biofilm producers. The major components of EOs were determined by gas chromatography-mass spectrometry (GC-MS). Biofilm formation of S. Typhimurium isolates was measured by crystal violet staining. Then, the effects of the EOs on the planktonic cell growth (using determination of minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC)), measurement of the synergistic effects of EOs (using checkerboard method), biofilm formation (using microtiter-plate test and scanning electron microscope (SEM)), and expression of QS and cellulose synthesis genes (using quantitative real-time PCR) were assessed. Finally, tetrazolium-based colorimetric (MTT) assay was used to examine EOs cytotoxicity on the Vero cell line. GC-MS analysis showed that terpineol, carene and pinene in C. cyminum EO and sulfur compounds in A. sativum EO were the major components of the plant extract. The Geometric mean of MIC values of the A. sativum and C. cyminum were 0.66 and 2.62 μL mL^-1, respectively. The geometric means of the fractional inhibitory concentration index (FICi) for both EOs were calculated as 1.05. The qPCR results showed that MIC/2 concentrations of both EOs significantly down-regulated of QS (sdiA and luxS) and cellulose synthesis (csgD and adrA) genes. Scanning electron microscopy showed the EOs reduced the amount of S. Typhimurium mature biofilm. In general, we showed that C. cyminum and A. sativum EOs can be considered as the potential agents against planktonic and biofilm form of S. Typhimurium without any concern of cytotoxic effect at 4 MIC concentrations on the eukaryotic Vero cells.

Keywords: Allium sativum; Biofilm; Cuminum cyminum; Quorum sensing; Salmonella typhimurium.